Effect of Incandescent Light on Collection of West Nile Virus Vectors Using CDC Miniature Light Traps in Northern Colorado.

To evaluate whether the presence of clear incandescent light was attractive or refractive to host-seeking mosquitoes in northern Colorado, a Bayesian hierarchical model was created to measure differences in trap effectiveness based on presence or absence of phototactic cues. A total of eight CDC miniature light traps (with and without light) were set weekly across four locations in northern Colorado between Weeks 23 and 32 of year 2020. Culex mosquitoes (Diptera: Culicidae) accounted for 81% of all collections in this study with two vectors of West Nile virus being represented. The probability of catching both Culex tarsalis Coquillett and Culex pipiens Linnaeus was reduced when traps were equipped with light, but the difference was not statistically significant for Culex tarsalis. The clear reduction in the number of Culex pipiens caught when these traps were equipped with light indicates negative phototactic behavior and underestimation with current surveillance strategies. Removal of light from these traps may aid our understanding of these species' distribution within the environment, improve collection efficiency, and help guide implementation of targeted control measures used in public health mosquito control.

Aedes aegypti miRNA-33 modulates permethrin induced toxicity by regulating VGSC transcripts.

Aedes aegypti is a major vector of Zika, dengue, and other arboviruses. Permethrin adulticidal spraying, which targets the voltage-gated sodium channel (VGSC), is commonly done to reduce local mosquito populations and protect humans from exposure to arbovirus pathogens transmitted by this dangerous pest. Permethrin resistance, however, is a growing problem and understanding its underlying molecular basis may identify avenues to combat it. We identified a single G:C polymorphism in pre-miR-33 that was genetically associated with permethrin resistance; resulting isoforms had structural differences that may affect DICER-1/pre-miRNA processing rates. We then assessed the effects of overexpression of pre-miR-33 isoforms on permethrin toxicological phenotypes, VGSC transcript abundance and protein levels for two genetically related mosquito strains. One strain had its naturally high permethrin resistance levels maintained by periodic treatment, and the other was released from selection. VGSC protein levels were lower in the permethrin resistant strain than in the related permethrin-susceptible strain. Overexpression of the G-pre-miR-33 isoform reduced VGSC expression levels in both strains. To further elucidate changes in gene expression associated with permethrin resistance, exome-capture gDNA deep sequencing, genetic association mapping and subsequent gene set enrichment analysis revealed that transport genes, in particular, were selected in resistant versus susceptible mosquitoes. Collectively, these data indicate that miR-33 regulates VGSC expression as part of a nuanced system of neuronal regulation that contributes to a network of heritable features determining permethrin resistance.

Vgsc-interacting proteins are genetically associated with pyrethroid resistance in Aedes aegypti.

Association mapping of factors that condition pyrethroid resistance in Aedes aegypti has consistently identified genes in multiple functional groups. Toward better understanding of the mechanisms involved, we examined high throughput sequencing data (HTS) from two Aedes aegypti aegypti collections from Merida, Yucatan, Mexico treated with either permethrin or deltamethrin. Exome capture enrichment for coding regions and the AaegL5 annotation were used to identify genes statistically associated with resistance. The frequencies of single nucleotide polymorphisms (SNPs) were compared between resistant and susceptible mosquito pools using a contingency χ2 analysis. The -log10(χ2 p value) was calculated at each SNP site, with a weighted average determined from all sites in each gene. Genes with -log10(χ2 p value) ≥ 4.0 and present among all 3 treatment groups were subjected to gene set enrichment analysis (GSEA). We found that several functional groups were enriched compared to all coding genes. These categories were transport, signal transduction and metabolism, in order from highest to lowest statistical significance. Strikingly, 21 genes with demonstrated association to synaptic function were identified. In the high association group (n = 1,053 genes), several genes were identified that also genetically or physically interact with the voltage-gated sodium channel (VGSC). These genes were eg., CHARLATAN (CHL), a transcriptional regulator, several ankyrin-domain proteins, PUMILIO (PUM), a translational repressor, and NEDD4 (E3 ubiquitin-protein ligase). There were 13 genes that ranked among the top 10%: these included VGSC; CINGULIN, a predicted neuronal gap junction protein, and the aedine ortholog of NERVY (NVY), a transcriptional regulator. Silencing of CHL and NVY followed by standard permethrin bottle bioassays validated their association with permethrin resistance. Importantly, VGSC levels were also reduced about 50% in chl- or nvy-dsRNA treated mosquitoes. These results are consistent with the contribution of a variety of neuronal pathways to pyrethroid resistance in Ae. aegypti.

Susceptibility and Vectorial Capacity of American Aedes albopictus and Aedes aegypti (Diptera: Culicidae) to American Zika Virus Strains.

The rapid expansion of Zika virus (ZIKV), following the recent outbreaks of Chikungunya virus, overwhelmed the public health infrastructure in many countries and alarmed many in the scientific community. Aedes aegypti (L.) (Diptera: Culicidae) and Aedes albopictus (Skuse) (Diptera: Culicidae) have previously been incriminated as the vectors of these pathogens in addition to dengue virus. In our study, we challenged low generation Ae. aegypti (Chiapas, Mexico) and Ae. albopictus (North Carolina, Mississippi), with three strains of ZIKV, Puerto Rico (GenBank: KU501215), Honduras (GenBank: KX694534), and Miami (GenBank: MF988743). Following an oral challenge with 107.5 PFU/ml of the Puerto Rico strain, we observed high infection and dissemination rates in both species (95%). We report estimated transmission rates for both species (74 and 33%, for Ae. aegypti (L.) (Diptera: Culicidae) and Ae. albopictus (Skuse) (Diptera: Culicidae), respectively), and the presence of a probable salivary gland barrier in Ae. albopictus to Zika virus. Finally, we calculated vectorial capacity for both species and found that Ae. albopictus had a slightly lower vectorial capacity when compared with Ae. aegypti.Second Language Abstract: La rápida expansión del virus Zika, poco después de las epidemias de chikungunya, rebaso la infraestructura de salud pública en muchos países y sorprendió a muchos en la comunidad científica. Notablemente, Aedes aegypti y Aedes albopictus transmiten estos patógenos además del virus del dengue. En este estudio se expusieron con tres cepas americanas de virus Zika a grupos de Aedes aegypti y Aedes albopictus de generación reciente. Encontramos altos porcentajes de infección y diseminación en ambas especies (95%). Se reporta, la transmisión viral en ambas especies (74 y 33%, para Aedes aegypti and Aedes albopictus, respectivamente) y una probable barrera a nivel de glándulas salivarías. Finalmente, calculamos la capacidad vectorial para ambas especies.

Alternative patterns of sex chromosome differentiation in Aedes aegypti (L).

BACKGROUND: Some populations of West African Aedes aegypti, the dengue and zika vector, are reproductively incompatible; our earlier study showed that divergence and rearrangements of genes on chromosome 1, which bears the sex locus (M), may be involved. We also previously described a proposed cryptic subspecies SenAae (PK10, Senegal) that had many more high inter-sex FST genes on chromosome 1 than did Ae.aegypti aegypti (Aaa, Pai Lom, Thailand). The current work more thoroughly explores the significance of those findings. RESULTS: Intersex standardized variance (FST) of single nucleotide polymorphisms (SNPs) was characterized from genomic exome capture libraries of both sexes in representative natural populations of Aaa and SenAae. Our goal was to identify SNPs that varied in frequency between males and females, and most were expected to occur on chromosome 1. Use of the assembled AaegL4 reference alleviated the previous problem of unmapped genes. Because the M locus gene nix was not captured and not present in AaegL4, the male-determining locus, per se, was not explored. Sex-associated genes were those with FST values ≥ 0.100 and/or with increased expected heterozygosity (H exp , one-sided T-test, p < 0.05) in males. There were 85 genes common to both collections with high inter-sex FST values; all genes but one were located on chromosome 1. Aaa showed the expected cluster of high inter-sex FST genes proximal to the M locus, whereas SenAae had inter-sex FST genes along the length of chromosome 1. In addition, the Aaa M-locus proximal region showed increased H exp levels in males, whereas SenAae did not. In SenAae, chromosomal rearrangements and subsequent suppressed recombination may have accelerated X-Y differentiation. CONCLUSIONS: The evidence presented here is consistent with differential evolution of proto-Y chromosomes in Aaa and SenAae.

Preliminary efficacy investigations of oral fipronil against Anopheles arabiensis when administered to Zebu cattle (Bos indicus) under field conditions.

Globally, malaria remains one of the most important vector-borne diseases despite the extensive use of vector control, including indoor residual spraying (IRS) and insecticide-treated nets (ITNs). These control methods target endophagic vectors, whereas some malaria vectors, such as Anopheles arabiensis, preferentially feed outdoors on cattle, making it a complicated vector to control using conventional strategies. Our study evaluated whether treating cattle with a capsule containing the active ingredient (AI) fipronil could reduce vector density and sporozoite rates, and alter blood feeding behavior, when applied in a small-scale field study. A pilot field study was carried out in the Samia District, Western Kenya, from May to July 2015. Four plots, each comprised of 50 huts used for sleeping, were randomly designated to serve as control or treatment. A week before cattle treatment, baseline mosquito collections were performed inside the houses using mechanical aspirators. Animals in the treatment (and buffer) were administered a single oral application of fipronil at ∼0.5mg/kg of body weight. Indoor mosquito collections were performed once a week for four weeks following treatment. Female mosquitoes were first identified morphologically to species complex, followed by PCR-based methods to obtain species identity, sporozoite presence, and the host source of the blood meal. All three species of anophelines found in the study area (An. gambiae s.s., An. arabiensis, An. funestus s.s.) were actively transmitting Plasmodium falciparum during the study period. The indoor resting density of An. arabiensis was significantly reduced in treatment plot one at three weeks post-treatment (T1) (efficacy=89%; T1 density=0.08, 95% credibility intervals [0.05, 0.10]; control plot density=0.78 [0.22, 0.29]) and at four weeks post-treatment (efficacy=64%; T1 density=0.16 [0.08, 0.14]; control plot density=0.48 [0.17, 0.22]). The reduction of An. arabiensis mosquitoes captured in the treatment plot two was higher: zero females were collected after treatment. The indoor resting density of An. gambiae s.s. was not significantly different between the treatment (T1, T2) and their corresponding control plots (C1, C2). An. funestus s.s. showed an increase in density over time. The results of this preliminary study suggest that treating cattle orally with fipronil, to target exophagic and zoophagic malaria vectors, could be a valuable control strategy to supplement existing vector control interventions which target endophilic anthropophilic species.

Evaluation of a topical formulation of eprinomectin against Anopheles arabiensis when administered to Zebu cattle (Bos indicus) under field conditions.

BACKGROUND: Although vector control strategies, such as insecticide-treated bed nets (ITNs) and indoor residual spraying (IRS) have been effective in Kenya the transmission of malaria continues to afflict western Kenya. This residual transmission is driven in part by Anopheles arabiensis, known for its opportunistic blood feeding behaviour and propensity to feed outdoors. The objective of this research was to evaluate the efficacy of the drug eprinomectin at reducing malaria vector density when applied to cattle (Bos indicus), the primary source of blood for An. arabiensis, under field conditions. METHODS: A pilot study was carried out in the Samia District of western Kenya from September to October of 2014. Treatment and control areas were randomly designated and comprised of 50 homes per study area. Before cattle treatments, baseline mosquito counts were performed after pyrethrum spray. Cows in the treatment area were administered topical applications of eprinomectin at 0.5 mg/kg once a week for two consecutive weeks. Mosquito collections were performed once each week for two weeks following the eprinomectin treatments. Mosquitoes were first identified morphologically and with molecular confirmation, then screened for sporozoite presence and host blood using PCR-based methods. RESULTS: The indoor resting density of An. arabiensis was significantly reduced by 38 % in the treatment area compared to the control area at one-week post-treatment (Control mean females per hut = 1.33 95 % CI [1.08, 1.64]; Treatment = 0.79 [0.56, 1.07]). An increase in the indoor resting density of Anopheles gambiae s.s. and Anopheles funestus s.s. was observed in the treatment area in the absence of An. arabiensis. At two weeks post-treatment, the total number of mosquitoes for any species per hut was not significantly different between the treatment and control areas. No change was observed in An. arabiensis host preference as a result of treatment. CONCLUSIONS: Systemic drugs may be an important tool by which to supplement existing vector control interventions by significantly impacting outdoor malaria transmission driven by An. arabiensis through the treatment of cattle.

WHATCH'EM: A Weather-Driven Energy Balance Model for Determining Water Height and Temperature in Container Habitats for Aedes aegypti.

The mosquito virus vector Aedes (Ae.) aegypti exploits a wide range of containers as sites for egg laying and development of the immature life stages, yet the approaches for modeling meteorologically sensitive container water dynamics have been limited. This study introduces the Water Height and Temperature in Container Habitats Energy Model (WHATCH'EM), a state-of-the-science, physically based energy balance model of water height and temperature in containers that may serve as development sites for mosquitoes. The authors employ WHATCH'EM to model container water dynamics in three cities along a climatic gradient in México ranging from sea level, where Ae. aegypti is highly abundant, to ~2100 m, where Ae. aegypti is rarely found. When compared with measurements from a 1-month field experiment in two of these cities during summer 2013, WHATCH'EM realistically simulates the daily mean and range of water temperature for a variety of containers. To examine container dynamics for an entire season, WHATCH'EM is also driven with field-derived meteorological data from May to September 2011 and evaluated for three commonly encountered container types. WHATCH'EM simulates the highly nonlinear manner in which air temperature, humidity, rainfall, clouds, and container characteristics (shape, size, and color) determine water temperature and height. Sunlight exposure, modulated by clouds and shading from nearby objects, plays a first-order role. In general, simulated water temperatures are higher for containers that are larger, darker, and receive more sunlight. WHATCH'EM simulations will be helpful in understanding the limiting meteorological and container-related factors for proliferation of Ae. aegypti and may be useful for informing weather-driven early warning systems for viruses transmitted by Ae. aegypti.

First report of L1014F kdr mutation in Culex quinquefasciatus in Mexico.

The L1014F mutation in the voltage-sodium channel gene has been associated with resistance to DDT and pyrethroids in various arthropod species including mosquitoes. We determined the frequency of the L1014F kdr mutation in 16 field populations of Culex quinquefasciatus from Northeastern Mexico collected between 2008 and 2013. The L1014F was present in all populations analyzed with the lowest frequency (3.33%) corresponding to the population from Monclova collected in 2012, and the highest frequency (63.63%) from the Monterrey population collected in 2012. The presence of a kdr mutation in populations of Cx. quinquefasciatus from northeastern Mexico provides evidence of pyrethroid resistance. This requires a special attention, considering that pyrethroid-based insecticides are commonly used in vector-control campaigns, especially against Aedes aegypti (L.).

Coevolution of the Ile1,016 and Cys1,534 Mutations in the Voltage Gated Sodium Channel Gene of Aedes aegypti in Mexico.

BACKGROUND: Worldwide the mosquito Aedes aegypti (L.) is the principal urban vector of dengue viruses. Currently 2.5 billion people are at risk for infection and reduction of Ae. aegypti populations is the most effective means to reduce the risk of transmission. Pyrethroids are used extensively for adult mosquito control, especially during dengue outbreaks. Pyrethroids promote activation and prolong the activation of the voltage gated sodium channel protein (VGSC) by interacting with two distinct pyrethroid receptor sites [1], formed by the interfaces of the transmembrane helix subunit 6 (S6) of domains II and III. Mutations of S6 in domains II and III synergize so that double mutants have higher pyrethroid resistance than mutants in either domain alone. Computer models predict an allosteric interaction between mutations in the two domains. In Ae. aegypti, a Ile1,016 mutation in the S6 of domain II was discovered in 2006 and found to be associated with pyrethroid resistance in field populations in Mexico. In 2010 a second mutation, Cys1,534 in the S6 of domain III was discovered and also found to be associated with pyrethroid resistance and correlated with the frequency of Ile1,016. METHODOLOGY/PRINCIPAL FINDINGS: A linkage disequilibrium analysis was performed on Ile1,016 and Cys1,534 in Ae. aegypti collected in Mexico from 2000-2012 to test for statistical associations between S6 in domains II and III in natural populations. We estimated the frequency of the four dilocus haplotypes in 1,016 and 1,534: Val1,016/Phe1,534 (susceptible), Val1,016/Cys1,534, Ile1,016/Phe1,534, and Ile1,016/Cys1,534 (resistant). The susceptible Val1,016/Phe1,534 haplotype went from near fixation to extinction and the resistant Ile1,016/Cys1,534 haplotype increased in all collections from a frequency close to zero to frequencies ranging from 0.5-0.9. The Val1,016/Cys1,534 haplotype increased in all collections until 2008 after which it began to decline as Ile1,016/Cys1,534 increased. However, the Ile1,016/Phe1,534 haplotype was rarely detected; it reached a frequency of only 0.09 in one collection and subsequently declined. CONCLUSION/SIGNIFICANCE: Pyrethroid resistance in the vgsc gene requires the sequential evolution of two mutations. The Ile1,016/Phe1,534 haplotype appears to have low fitness suggesting that Ile1,016 was unlikely to have evolved independently. Instead the Cys1,534 mutation evolved first but conferred only a low level of resistance. Ile1,016 in S6 of domain II then arose from the Val1,016/Cys1,534 haplotype and was rapidly selected because double mutants confer higher pyrethroid resistance. This pattern suggests that knowledge of the frequencies of mutations in both S6 in domains II and III are important to predict the potential of a population to evolve kdr. Susceptible populations with high Val1,016/Cys1,534 frequencies are at high risk for kdr evolution, whereas susceptible populations without either mutation are less likely to evolve high levels of kdr, at least over a 10 year period.

Age and prior blood feeding of Anopheles gambiae influences their susceptibility and gene expression patterns to ivermectin-containing blood meals.

BACKGROUND: Ivermectin has been proposed as a novel malaria transmission control tool based on its insecticidal properties and unique route of acquisition through human blood. To maximize ivermectin's effect and identify potential resistance/tolerance mechanisms, it is important to understand its effect on mosquito physiology and potential to shift mosquito population age-structure. We therefore investigated ivermectin susceptibility and gene expression changes in several age groups of female Anopheles gambiae mosquitoes. METHODS: The effect of aging on ivermectin susceptibility was analyzed in three age groups (2, 6, and 14-days) of colonized female Anopheles gambiaemosquitoes using standard survivorship assays. Gene expression patterns were then analyzed by transcriptome sequencing on an Illumina HiSeq 2500 platform. RT-qPCR was used to validate transcriptional changes and also to examine expression in a different, colonized strain and in wild mosquitoes, both of which blood fed naturally on an ivermectin-treated person. RESULTS: Mosquitoes of different ages and blood meal history died at different frequencies after ingesting ivermectin. Mortality was lowest in 2-day old mosquitoes exposed on their first blood meal and highest in 6-day old mosquitoes exposed on their second blood meal. Twenty-four hours following ivermectin ingestion, 101 and 187 genes were differentially-expressed relative to control blood-fed, in 2 and 6-day groups, respectively. Transcription patterns of select genes were similar in membrane-fed, colonized, and naturally-fed wild vectors. Transcripts from several unexpected functional classes were highly up-regulated, including Niemann-Pick Type C (NPC) genes, peritrophic matrix-associated genes, and immune-response genes, and these exhibited different transcription patterns between age groups, which may explain the observed susceptibility differences. Niemann-Pick Type 2 genes were the most highly up-regulated transcripts after ivermectin ingestion (up to 160 fold) and comparing phylogeny to transcriptional patterns revealed that NPCs have rapidly evolved and separate members respond to either blood meals or to ivermectin. CONCLUSION: We present evidence of increased ivermectin susceptibility in older An. gambiae mosquitoes that had previously bloodfed. Differential expression analysis suggests complex midgut interactions resulting from ivermectin ingestion that likely involve blood meal digestion physiological responses, midgut microflora, and innate immune responses. Thus, the transcription of certain gene families is consistently affected by ivermectin ingestion, and may provide important clues to ivermectin's broad effects on malaria vectors. These findings contribute to the growing understanding of ivermectin's potential as a transmission control tool.

Assessing Insecticide Susceptibility of Laboratory Lutzomyia longipalpis and Phlebotomus papatasi Sand Flies (Diptera: Psychodidae: Phlebotominae).

Chemical insecticides are effective for controlling Lutzomyia and Phlebotomus sand fly (Diptera: Psychodidae) vectors of Leishmania parasites. However, repeated use of certain insecticides has led to tolerance and resistance. The objective of this study was to determine lethal concentrations (LCs) and lethal exposure times (LTs) to assess levels of susceptibility of laboratory Lutzomyia longipalpis (Lutz and Nieva) and Phlebotomus papatasi (Scopoli) to 10 insecticides using a modified version of the World Health Organization (WHO) exposure kit assay and Centers for Disease Control and Prevention (CDC) bottle bioassay. Sand flies were exposed to insecticides coated on the interior of 0.5-gallon and 1,000-ml glass bottles. Following exposure, the flies were allowed to recover for 24 h, after which mortality was recorded. From dose-response survival curves for L. longipalpis and P. papatasi generated with the QCal software, LCs causing 50, 90, and 95% mortality were determined for each insecticide. The LCs and LTs from this study will be useful as baseline reference points for future studies using the CDC bottle bioassays to assess insecticide susceptibility of sand fly populations in the field. There is a need for a larger repository of sand fly insecticide susceptibility data from the CDC bottle bioassays, including a range of LCs and LTs for more sand fly species with more insecticides. Such a repository would be a valuable tool for vector management.

Local evolution of pyrethroid resistance offsets gene flow among Aedes aegypti collections in Yucatan State, Mexico.

The mosquito Aedes aegypti is the major vector of the four serotypes of dengue virus (DENV1-4). Previous studies have shown that Ae. aegypti in Mexico have a high effective migration rate and that gene flow occurs among populations that are up to 150 km apart. Since 2000, pyrethroids have been widely used for suppression of Ae. aegypti in cities in Mexico. In Yucatan State in particular, pyrethroids have been applied in and around dengue case households creating an opportunity for local selection and evolution of resistance. Herein, we test for evidence of local adaptation by comparing patterns of variation among 27 Ae. aegypti collections at 13 single nucleotide polymorphisms (SNPs): two in the voltage-gated sodium channel gene para known to confer knockdown resistance, three in detoxification genes previously associated with pyrethroid resistance, and eight in putatively neutral loci. The SNPs in para varied greatly in frequency among collections, whereas SNPs at the remaining 11 loci showed little variation supporting previous evidence for extensive local gene flow. Among Ae. aegypti in Yucatan State, Mexico, local adaptation to pyrethroids appears to offset the homogenizing effects of gene flow.

Intra-annual changes in abundance of Aedes (Stegomyia) aegypti and Aedes (Ochlerotatus) epactius (Diptera: Culicidae) in high-elevation communities in Mexico.

We examined temporal changes in the abundance of the mosquitoes Aedes (Stegomyia) aegypti (L.) and Aedes (Ochlerotatus) epactius Dyar & Knab from June to October 2012 in one reference community at lower elevation (Rio Blanco; approximately 1,270 m) and three high-elevation communities (Acultzingo, Maltrata, and Puebla City; 1,670-2,150 m) in Veracruz and Puebla States, México. The combination of surveys for pupae in water-filled containers and trapping of adults, using BG-Sentinel traps baited with the BG-Lure, corroborated previous data from 2011 showing that Ae. aegypti is present at low abundance up to 2,150 m in this part of México. Data for Ae. aegypti adults captured through repeated trapping in fixed sites in Acultzingo--the highest elevation community (approximately 1,670 m) from which the temporal intra-annual abundance pattern for Ae. aegypti has been described--showed a gradual increase from low numbers in June to a peak occurrence in late August, and thereafter declining numbers in September. Ae. epactius adults were collected repeatedly in BG-Sentinel traps in all four study communities; this is the first recorded collection of this species with a trap aiming specifically to collect human-biting mosquitoes. We also present the first description of the temporal abundance pattern for Ae. epactius across an elevation gradient: peak abundance was reached in mid-July in the lowest elevation community (Rio Blanco) but not until mid-September in the highest elevation one (Puebla City). Finally, we present data for meteorological conditions (mean temperature and rainfall) in the examined communities during the study period, and for a cumulative measure of the abundance of adults over the full sampling period.

The impact of temperature on the bionomics of Aedes (Stegomyia) aegypti, with special reference to the cool geographic range margins.

The mosquito Aedes (Stegomyia) aegypti (L.), which occurs widely in the subtropics and tropics, is the primary urban vector of dengue and yellow fever viruses, and an important vector of chikungunya virus. There is substantial interest in how climate change may impact the bionomics and pathogen transmission potential of this mosquito. This Forum article focuses specifically on the effects of temperature on the bionomics of Ae. aegypti, with special emphasis on the cool geographic range margins where future rising temperatures could facilitate population growth. Key aims are to: 1) broadly define intra-annual (seasonal) patterns of occurrence and abundance of Ae. aegypti, and their relation to climate conditions; 2) synthesize the existing quantitative knowledge of how temperature impacts the bionomics of different life stages of Ae. aegypti; 3) better define the temperature ranges for which existing population dynamics models for Ae. aegypti are likely to produce robust predictions; 4) explore potential impacts of climate warming on human risk for exposure to Ae. aegypti at its cool range margins; and 5) identify knowledge or data gaps that hinder our ability to predict risk of human exposure to Ae. aegypti at the cool margins of its geographic range now and in the future. We first outline basic scenarios for intra-annual occurrence and abundance patterns for Ae. aegypti, and then show that these scenarios segregate with regard to climate conditions in selected cities where they occur. We then review how near-constant and intentionally fluctuating temperatures impact development times and survival of eggs and immatures. A subset of data, generated in controlled experimental studies, from the published literature is used to plot development rates and survival of eggs, larvae, and pupae in relation to water temperature. The general shape of the relationship between water temperature and development rate is similar for eggs, larvae, and pupae. Once the lower developmental zero temperature (10-14 degrees C) is exceeded, there is a near-linear relationship up to 30 degrees C. Above this temperature, the development rate is relatively stable or even decreases slightly before falling dramatically near the upper developmental zero temperature, which occurs at -38-42 degrees C. Based on life stage-specific linear relationships between water temperature and development rate in the 15-28 degrees C range, the lower developmental zero temperature is estimated to be 14.0 degrees C for eggs, 11.8 degrees C for larvae, and 10.3 degrees C for pupae. We further conclude that available population dynamics models for Ae. aegypti, such as CIMSiM and Skeeter Buster, likely produce robust predictions based on water temperatures in the 16-35 degrees C range, which includes the geographic areas where Ae. aegypti and its associated pathogens present the greatest threat to human health, but that they may be less reliable in cool range margins where water temperatures regularly fall below 15 degrees C. Finally, we identify knowledge or data gaps that hinder our ability to predict risk of human exposure to Ae. aegypti at the cool margins of its range, now and in the future, based on impacts on mosquito population dynamics of temperature and other important factors, such as water nutrient content, larval density, presence of biological competitors, and human behavior.

Cell phone-based system (Chaak) for surveillance of immatures of dengue virus mosquito vectors.

Capture of surveillance data on mobile devices and rapid transfer of such data from these devices into an electronic database or data management and decision support systems promote timely data analyses and public health response during disease outbreaks. Mobile data capture is used increasingly for malaria surveillance and holds great promise for surveillance of other neglected tropical diseases. We focused on mosquito-borne dengue, with the primary aims of: 1) developing and field-testing a cell phone-based system (called Chaak) for capture of data relating to the surveillance of the mosquito immature stages, and 2) assessing, in the dengue endemic setting of Mérida, Mexico, the cost-effectiveness of this new technology versus paper-based data collection. Chaak includes a desktop component, where a manager selects premises to be surveyed for mosquito immatures, and a cell phone component, where the surveyor receives the assigned tasks and captures the data. Data collected on the cell phone can be transferred to a central database through different modes of transmission, including near-real time where data are transferred immediately (e.g., over the Internet) or by first storing data on the cell phone for future transmission. Spatial data are handled in a novel, semantically driven, geographic information system. Compared with a pen-and-paper-based method, use of Chaak improved the accuracy and increased the speed of data transcription into an electronic database. The cost-effectiveness of using the Chaak system will depend largely on the up-front cost of purchasing cell phones and the recurring cost of data transfer over a cellular network.

Towards a Casa Segura: a consumer product study of the effect of insecticide-treated curtains on Aedes aegypti and dengue virus infections in the home.

The home, or domicile, is the principal environment for transmission of dengue virus (DENV) between humans and mosquito vectors. Community-wide distribution of insecticide-treated curtains (ITCs), mimicking vector control program-driven interventions, has shown promise to reduce DENV infections. We conducted a Casa Segura consumer product intervention study in Mérida, Mexico to determine the potential to reduce intradomicillary DENV transmission through ITC use in individual homes. Dengue virus infections in mosquitoes and in humans were reduced in homes with ITCs in one of two study subareas. Overall, ITCs reduced intradomicillary DENV transmission; ITC homes were significantly less likely to experience multiple DENV infections in humans than NTC homes. Dengue virus-infected Aedes aegypti females were reduced within the ITC homes where curtain use was highest. Some homes yielded up to nine infected Ae. aegypti females. This study provides insights regarding best practices for Casa Segura interventions to protect homes from intradomicillary DENV transmission.

Ontology for vector surveillance and management.

Ontologies, which are made up by standardized and defined controlled vocabulary terms and their interrelationships, are comprehensive and readily searchable repositories for knowledge in a given domain. The Open Biomedical Ontologies (OBO) Foundry was initiated in 2001 with the aims of becoming an "umbrella" for life-science ontologies and promoting the use of ontology development best practices. A software application (OBO-Edit; *.obo file format) was developed to facilitate ontology development and editing. The OBO Foundry now comprises over 100 ontologies and candidate ontologies, including the NCBI organismal classification ontology (NCBITaxon), the Mosquito Insecticide Resistance Ontology (MIRO), the Infectious Disease Ontology (IDO), the IDOMAL malaria ontology, and ontologies for mosquito gross anatomy and tick gross anatomy. We previously developed a disease data management system for dengue and malaria control programs, which incorporated a set of information trees built upon ontological principles, including a "term tree" to promote the use of standardized terms. In the course of doing so, we realized that there were substantial gaps in existing ontologies with regards to concepts, processes, and, especially, physical entities (e.g., vector species, pathogen species, and vector surveillance and management equipment) in the domain of surveillance and management of vectors and vector-borne pathogens. We therefore produced an ontology for vector surveillance and management, focusing on arthropod vectors and vector-borne pathogens with relevance to humans or domestic animals, and with special emphasis on content to support operational activities through inclusion in databases, data management systems, or decision support systems. The Vector Surveillance and Management Ontology (VSMO) includes >2,200 unique terms, of which the vast majority (>80%) were newly generated during the development of this ontology. One core feature of the VSMO is the linkage, through the has vector relation, of arthropod species to the pathogenic microorganisms for which they serve as biological vectors. We also recognized and addressed a potential roadblock for use of the VSMO by the vector-borne disease community: the difficulty in extracting information from OBO-Edit ontology files (*.obo files) and exporting the information to other file formats. A novel ontology explorer tool was developed to facilitate extraction and export of information from the VSMO*.obo file into lists of terms and their associated unique IDs in *.txt or *.csv file formats. These lists can then be imported into a database or data management system for use as select lists with predefined terms. This is an important step to ensure that the knowledge contained in our ontology can be put into practical use.

Aedes (Ochlerotatus) epactius along an elevation and climate gradient in Veracruz and Puebla States, México.

We report on the collection ofimmatures of Aedes (Ochlerotatus) epactius Dyar & Knab from artificial containers during July through September 2011 in 12 communities located along an elevation and climate gradient extending from sea level in Veracruz State to high elevations (>2,000 m) in Veracruz and Puebla States, México. Ae. epactius was collected from 11 of the 12 study communities; the lone exception was the highest elevation community along the transect (>2,400 m). This mosquito species was thus encountered at elevations ranging from near sea level in Veracruz City on the Gulf of México to above 2,100 m in Puebla City in the central highlands. Collection sites included the city of C6rdoba, located at approximately 850 m, from which some of the first described specimens of Ae. epactius were collected in 1908. Estimates for percentage of premises in each community with Ae. epactius pupae present, and abundance of Ae. epactius pupae on the study premises, suggest that along the transect in central México, the mosquito is present but rare at sea level, most abundant at mid-range elevations from 1,250-1,750 m and then decreases in abundance above 1,800 m. Statistically significant parabolic relationships were found between percentage of premises with Ae. epactius pupae present and average minimum daily temperature, cumulative growing degree-days, and rainfall. We recorded Ae. epactius immatures from a wide range of container types including cement water tanks, barrels/ drums, tires, large earthen jars, small discarded containers, buckets, cement water troughs, flower pots, cement water cisterns, and larger discarded containers. There were 45 documented instances of co-occurrence of Ae. epactius and Aedes aegypti (L.) immatures in individual containers.

Aedes (Stegomyia) aegypti and Aedes (Howardina) cozumelensis in Yucatán State, México, with a summary of published collection records for Ae. cozumelensis.

We collected mosquito immatures from artificial containers during 2010-2011 from 26 communities, ranging in size from small rural communities to large urban centers, located in different parts of Yucatán State in southeastern México. The arbovirus vector Aedes (Stegomyia) aegypti was collected from all 26 examined communities, and nine of the communities also yielded another container-inhabiting Aedes mosquito: Aedes (Howardina) cozumelensis. The communities from which Ae. cozumelensis were collected were all small rural communities (<6,000 inhabitants) in the north-central part of Yucatán State. These new collection records for Ae. cozumelensis demonstrate that this mosquito has a far broader geographic range in the Yucatán Peninsula than previously known. Ae. cozumelensis immatures were collected from both residential premises and cemeteries, with specimens recovered from rock holes as well as various artificial containers including metal cans, flower vases, buckets, tires, and a water storage tank. The co-occurrence with Ae. aegypti in small rural communities poses intriguing questions regarding linkages between these mosquitoes, including the potential for direct competition for larval development sites. Additional studies are needed to determine how commonly Ae. cozumelensis feeds on human blood and whether it is naturally infected with arboviruses or other pathogens of medical or veterinary importance. We also summarize the published records for Ae. cozumelensis, which are restricted to collections from México's Yucatán Peninsula and Belize, and uniformly represent geographic locations where Ae. aegypti can be expected to occur.